DNA Methylation Based Biomarkers and Epigenetic Regulation in IBS

 描述（由申请人提供）：肠易激综合征（IBS）是一种慢性胃肠道疾病，在美国影响高达15%的成年人，主要是女性。症状包括与腹泻、便秘或两者相关的慢性或复发性腹痛。它被认为是一种压力敏感性疾病，与脑肠道相互作用的改变有关。目前缺乏能够可靠诊断IBS或监测治疗反应的生物标志物。目前的建议是基于我们的初步数据，即表观遗传标记，即DNA甲基化，可以区分IBS患者和健康个体。DNA甲基化是控制脊椎动物基因功能的关键表观遗传机制。它已成为一种主要机制，将基因-环境相互作用与长期行为发育联系起来，特别是在IBS等复杂疾病中。我们对有限数量的受试者（IBS：N=12，健康对照：N = 12）外周血单核细胞（PBMC）全基因组DNA甲基化的初步数据确定了一组位于基因上的表观遗传标记，包括SSPO，RNF 39，GSTM 1，GSTM 5，TPPP和SNCAIP，可以潜在地区分IBS患者和健康对照。基因本体分析表明，丰富的基因参与神经肽途径。我们使用亚硫酸氢盐测序验证了这些基因中CpG位点的差异甲基化。然而，这些发现需要在更大的队列中进行复制和验证。因此，本文提出的研究旨在确定用于准确诊断IBS的稳健的表观遗传生物标志物，并获得对这种疾病发病机制的重要见解。我们的目标是：1）使用IBS中外周血单核细胞（PBMC）中的全基因组DNA甲基化谱鉴定基于DNA甲基化的生物标志物。2)研究IBS患者和HC结肠组织中全基因组甲基化和表达差异。在目标1中，我们将在一个更大的、独立的IBS患者（N=108）和对照组（N=36）队列中进行全基因组DNA甲基化分析，使用可用于IBS常规诊断的负担得起的技术（MethyLight PCR）复制样本子集中选定标记物的差异甲基化。在目标2中，我们提出了从IBS患者（N=108）和对照组（N=36）收集的库存结肠粘膜活检组织的全基因组DNA甲基化和基因表达谱，以研究表观遗传学失调的基因和相关通路，这可以为IBS的病理生理学提供重要的见解。

 DESCRIPTION (provided by applicant): Irritable bowel syndrome (IBS) is a chronic gastrointestinal disorder that affects up to 15% of adults, predominantly women, in the United States. Symptoms include chronic or recurrent abdominal pain associated with diarrhea, constipation or both. It is considered a stress-sensitive disorder that is associated with altered brain-gut interactions. Biomarkers that can reliably diagnose IBS or monitor treatment response are currently lacking. The current proposal is based on our preliminary data that epigenetic markers, namely DNA methylation, can distinguish IBS patients from healthy individuals. DNA methylation is a key epigenetic mechanism that governs vertebrate gene function. It has emerged as a leading mechanism linking gene- environment interactions to long-term behavioral development, particularly in complex disorders such as IBS. Our preliminary data on genome-wide DNA methylation for peripheral blood mononuclear cells (PBMCs) in a limited number of subjects (IBS: N=12, healthy controls: N=12) identified a set of epigenetic markers located on genes including SSPO, RNF39, GSTM1, GSTM5, TPPP and SNCAIP that can potentially distinguish IBS patients from healthy controls. Gene ontology analysis showed an enrichment of genes involved in neuropeptide pathways. We validated the differential methylation of CpG sites in these genes using bisulphite sequencing. However, these findings need to be replicated and validated in a larger cohort. Therefore, the studies proposed here are intended to identify robust epigenetic biomarkers for an accurate diagnosis of IBS and to gain critical insights into the pathogenesis of this disorder. Our aims are to: 1) Identify DNA methylation based biomarkers using genome-wide DNA methylation profiling in peripheral blood mononuclear cells (PBMCs) in IBS. 2) Study genome-wide methylation and expression differences in colon tissue of IBS patients and HCs. In Aim 1, we will conduct a genome-wide DNA methylation profiling in a larger, independent cohort of IBS patients (N=108) and controls (N=36) replicate the differential methylation of selected markers in a subset of samples using an affordable technique (MethyLight PCR) that can be employed for the routine diagnosis of IBS. In Aim 2, we propose genome-wide DNA methylation and gene expression profiling of banked colonic mucosal biopsies collected from IBS patients (N=108) and controls (N=36) to investigate epigenetically deregulated genes and associated pathways, which can give important insights on pathophysiology of IBS.